Defibrillation is routinely used in patients to interrupt ventricular fibrillation, yet there is a need to improve its efficacy and reduce side effects. To achieve these goals, a better understanding of the interaction between defibrillation shocks and cardiac tissue is needed. This application will address two fundamental questions: (1) how electrical field produces changes of transmembrane potential (deltaVm), which are necessary for defibrillation; (2) how strong defibrillation shocks induce arrhythmias, that can cause defibrillation failure. To achieve a deeper mechanistic understanding of these events, we will monitor spatio-temporal dynamics of Vm and Ca 2+ changes during shocks using novel techniques for simultaneous optical imaging of Vm and Cai2+ and ratiometric imaging of Vm. Studies will be performed in two experimental models that offer unique advantages for studying defibrillation mechanisms: patterned growth cell cultures and coronary perfused wedges of pig ventricles. Using 2-dimensional cell cultures will allow for precise control of tissue structure, electrical field and cell environment. Using coronary perfused wedges of ventricular myocardium will allow to obtain information on intramural shock-induced changes of Vm. The project will have three Specific Aims. 1) To determine the mechanisms of nonlinear shock-induced deltaVm. Recent optical mapping studies showed that defibrillation shocks induce complex nonlinear deltaVm. The roles of ionic currents and membrane electroporation in nonlinear deltaVm responses will be determined in cell cultures using simultaneous Vm/Cai2+ mapping and ionic channel blockers. 2) To determine the mechanism of arrhythmias induced by strong shocks. Using patterned growth cell cultures we will determine the type of deltaVm causing post-shock arrhythmias, the roles of Cai2+ overload and Vm depolarization, modulation of arrhythmia initiation by tissue structure, hyperkalemia and shock waveform. 3) To determine the distribution and magnitude of intramural deltaVm. Presently, no data are available on deltaVm in the intramural layers of myocardium during shocks. We will use optical mapping of Vm to measure the distributions of deltaVm in intramural layers of coronary-perfused wedges of pig ventricles and the effects of deltaVm on the duration of intramural action potentials.